10
From DNA to Protein: Gene
Expression
Chapter 10 From DNA to Protein: Gene Expression
Key Concepts
• 10.1 Genetics Shows That Genes Code
for Proteins
• 10.2 DNA Expression Begins with Its
Transcription to RNA
• 10.3 The Genetic Code in RNA Is
Translated into the Amino Acid Sequences
of Proteins
Chapter 10 From DNA to Protein: Gene Expression
• 10.4 Translation of the Genetic Code is
Mediated by tRNA and Ribosomes
• 10.5 Proteins Are Modified after
Translation
Chapter 10 Opening Question
How do antibiotics such as tetracycline
target bacterial protein synthesis?
Concept 10.1 Genetics Shows That Genes Code for Proteins
Identification of a gene product as a protein
began with a mutation.
Garrod saw a disease phenotype—
alkaptonuria—occurring in children who
shared more alleles as first cousins.
A substance in their blood (HA)
accumulated—was not catalyzed—and the
gene for the enzyme was mutated.
Garrod correlated one gene to one enzyme.
Concept 10.1 Genetics Shows That Genes Code for Proteins
Phenylketonuria is another genetic disease
that involves this pathway.
The enzyme that converts phenylalanine to
tyrosine is nonfunctional.
Untreated, it can lead to mental retardation,
but is easily detected in newborns.
Figure 10.1 Metabolic Diseases and Enzymes (Part 1)
Figure 10.1 Metabolic Diseases and Enzymes (Part 2)
Concept 10.1 Genetics Shows That Genes Code for Proteins
Phenotypic expression of alkapatonuria and
phenylketonuria led to the one gene–one
protein hypothesis.
A mutant phenotype arises from a change in
the protein’s amino acid sequence.
However, the one gene–one protein
hypothesis proved too simple in studies of
human mutations.
Concept 10.1 Genetics Shows That Genes Code for Proteins
The gene–enzyme relationship has since
been revised to the one gene–one
polypeptide relationship.
Example: In hemoglobin, each polypeptide
chain is specified by a separate gene.
Other genes code for RNA but are not
translated to polypeptides; some genes
are involved in controlling other genes.
Figure 10.2 Gene Mutations and Amino Acid Changes
Concept 10.1 Genetics Shows That Genes Code for Proteins
Molecular biology is the study of nucleic
acids and proteins,and often focuses on
gene expression.
Gene expression to form a specific
polypeptide occurs in two steps:
• Transcription—copies information from a
DNA sequence (a gene) to a
complementary RNA sequence
• Translation—converts RNA sequence to
amino acid sequence of a polypeptide
Concept 10.1 Genetics Shows That Genes Code for Proteins
Roles of three kinds of RNA in protein
synthesis:
• Messenger RNA (mRNA) and
transcription—carries copy of a DNA
sequence to the site of protein synthesis at
the ribosome
• Ribosomal RNA (rRNA) and translation—
catalyzes peptide bonds between amino
acids
• Transfer RNA (tRNA) mediates between
mRNA and protein—carries amino acids
for polypeptide assembly
Figure 10.3 From Gene to Protein
Concept 10.2 DNA Expression Begins with Its Transcription to
RNA
Transcription—the formation of a specific
RNA sequence from a specific DNA
sequence—requires some components:
• A DNA template for base pairings—one of
the two strands of DNA
• Nucleoside triphosphates
(ATP,GTP,CTP,UTP) as substrates
• An RNA polymerase enzyme
Concept 10.2 DNA Expression Begins with Its Transcription to
RNA
Besides mRNAs, other types of RNA are
produced by transcription:
• tRNA
• rRNA
• Small nuclear RNAs
• microRNAs
RNAs may have other functions in the cell
besides protein synthesis.
Concept 10.2 DNA Expression Begins with Its Transcription to
RNA
RNA polymerases catalyze synthesis of
RNA from the DNA template.
RNA polymerases are processive—a single
enzyme-template binding results in
polymerization of hundreds of RNA bases.
Unlike DNA polymerases, RNA
polymerases do not need primers.
Figure 10.4 RNA Polymerase
Concept 10.2 DNA Expression Begins with Its Transcription to
RNA
Transcription occurs in three phases:
• Initiation
• Elongation
• Termination
Concept 10.2 DNA Expression Begins with Its Transcription to
RNA
Initiation requires a promoter—a special
sequence of DNA.
RNA polymerase binds to the promoter.
Promoter tells RNA polymerase two things:
• Where to start transcription
• Which strand of DNA to transcribe
Part of each promoter is the transcription
initiation site.
Figure 10.5 DNA Is Transcribed to Form RNA (Part 1)
Figure 10.5 DNA Is Transcribed to Form RNA (Part 2)
Concept 10.2 DNA Expression Begins with Its Transcription to
RNA
Elongation: RNA polymerase unwinds DNA
about 13 base pairs at a time; reads
template in 3′-to-5′ direction.
RNA polymerase adds nucleotides to the 3′
end of the new strand.
The first nucleotide in the new RNA forms
its 5′ end and the RNA transcript is
antiparallel to the DNA template strand.
RNA polymerases can proofread, but allow
more mistakes.
Figure 10.5 DNA Is Transcribed to Form RNA (Part 3)
Concept 10.2 DNA Expression Begins with Its Transcription to
RNA
Termination is specified by a specific DNA
base sequence.
Mechanisms of termination are complex and
varied.
For some genes, the transcript falls away
from the DNA template and RNA
polymerase—for others a helper protein
pulls it away.
Figure 10.5 DNA Is Transcribed to Form RNA (Part 4)
Concept 10.2 DNA Expression Begins with Its Transcription to
RNA
Coding regions are sequences of a DNA
molecule that are expressed as proteins.
Eukaryotic genes may have noncoding
sequences—introns (intervening
regions).
The coding sequences are exons
(expressed regions).
Introns and exons appear in the primary
mRNA transcript—pre-mRNA; introns are
removed from the final mRNA.
Figure 10.6 Transcription of a Eukaryotic Gene (Part 1)
Figure 10.6 Transcription of a Eukaryotic Gene (Part 2)
Concept 10.2 DNA Expression Begins with Its Transcription to
RNA
Nucleic acid hybridization reveals introns.
Target DNA is denatured, then incubated
with a probe—a nucleic acid strand from
another source.
If the probe has a complementary
sequence, a probe–target double helix—
called a hybrid—forms.
Figure 10.7 Nucleic Acid Hybridization
Concept 10.2 DNA Expression Begins with Its Transcription to
RNA
Researchers using mature mRNA as the
probe saw loops where base pairing did
not occur in the DNA–RNA hybrid.
If pre-mRNA was used, the result was a
linear matchup—complete hybridization.
Introns were a part of the pre-mRNA, but
were removed before primary mRNA was
made.
Figure 10.8 Demonstrating the Existence of Introns
Concept 10.2 DNA Expression Begins with Its Transcription to
RNA
Introns interrupt, but do not scramble, the
DNA sequence that encodes a
polypeptide.
Sometimes, the separated exons code for
different domains (functional regions) of
the protein.
Concept 10.2 DNA Expression Begins with Its Transcription to
RNA
RNA splicing removes introns and splices
exons together.
Newly transcribed pre-mRNA is bound at
ends by snRNPs—small nuclear
ribonucleoprotein particles.
Consensus sequences are short
sequences between exons and introns,
bound by snRNPs.
Concept 10.2 DNA Expression Begins with Its Transcription to
RNA
Besides the snRNPs, other proteins are
added to form an RNA–protein complex,
the spliceosome.
The complex cuts pre-mRNA, releases
introns, and splices exons together to
produce mature mRNA.
Figure 10.9 The Spliceosome: An RNA Splicing Machine
Concept 10.2 DNA Expression Begins with Its Transcription to
RNA
In the disease β-thalassemia, a mutation
may occur at an intron consensus
sequence in the β-globin gene—the premRNA can not be spliced correctly.
Non-functional β-globin mRNA is produced,
which shows how mutations are used to
elucidate cause-and-effect relationships.
Alternative splicing results in different
mRNAs and different polypeptides from a
single gene.
Concept 10.2 DNA Expression Begins with Its Transcription to
RNA
While the pre-mRNA is in the nucleus it
undergoes two processing steps:
A 5′ cap (or G cap) is added to the 5′ end as
it is transcribed and facilitates binding and
prevents breakdown by enzymes.
A poly A tail is added to the 3′ end at the
end of transcription and assists in export
from the nucleus and aids stability.
Concept 10.3 The Genetic Code in RNA Is Translated into the
Amino Acid Sequences of Proteins
The genetic code—specifies which amino
acids will be used to build a protein
Codon—a sequence of three bases; each
codon specifies a particular amino acid
Start codon—AUG—initiation signal for
translation
Stop codons—UAA, UAG, UGA—stop
translation and polypeptide is released
Figure 10.10 Deciphering the Genetic Code (Part 1)
Figure 10.11 The Genetic Code
Concept 10.3 The Genetic Code in RNA Is Translated into the
Amino Acid Sequences of Proteins
For most amino acids, there is more than
one codon; the genetic code is redundant.
The genetic code is not ambiguous—each
codon specifies only one amino acid.
The genetic code is nearly universal: the
codons that specify amino acids are the
same in all organisms.
Exceptions: Within mitochondria,
chloroplasts, and some protists, there are
differences.
Concept 10.3 The Genetic Code in RNA Is Translated into the
Amino Acid Sequences of Proteins
This common genetic code is a common
language for evolution.
The code is ancient and has remained intact
throughout evolution.
The common code also facilitates genetic
engineering.
Concept 10.3 The Genetic Code in RNA Is Translated into the
Amino Acid Sequences of Proteins
Mutations can also be defined in terms of
their effects on polypeptide sequences.
Silent mutations have no effect on amino
acids—often found in noncoding regions of
DNA.
A base substitution does not always affect
amino acid sequence, which may be
repaired in translation.
Figure 10.12 Mutations (Part 1)
Concept 10.3 The Genetic Code in RNA Is Translated into the
Amino Acid Sequences of Proteins
Missense mutations are substitutions by one
amino acid for another in a protein.
Example: Sickle-cell disease—allele differs
from normal by one base pair
Missense mutations may result in a
defective protein, reduced protein
efficiency, or even a gain of function as in
the TP53 gene.
Figure 10.12 Mutations (Part 2)
Concept 10.3 The Genetic Code in RNA Is Translated into the
Amino Acid Sequences of Proteins
Nonsense mutations involve a base
substitution that causes a stop codon to
form somewhere in the mRNA.
This results in a shortened protein, which is
usually not functional—if near the 3' end it
may have no effect.
Figure 10.12 Mutations (Part 3)
Concept 10.3 The Genetic Code in RNA Is Translated into the
Amino Acid Sequences of Proteins
Frame-shift mutations are insertions or
deletions of bases in DNA.
These mutations interfere with translation
and shift the “reading-frame.”
Nonfunctional proteins are produced.
Figure 10.12 Mutations (Part 4)
Concept 10.4 Translation of the Genetic Code Is Mediated by
tRNA and Ribosomes
tRNA links information in mRNA codons
with specific amino acids.
For each amino acid, there is a specific type
or “species” of tRNA.
Two key events to ensure that the protein
made is the one specified by the mRNA:
• tRNAs must read mRNA codons correctly.
• tRNAs must deliver amino acids
corresponding to each codon.
Concept 10.4 Translation of the Genetic Code Is Mediated by
tRNA and Ribosomes
Each tRNA has three functions, made
possible by its structure and base
sequence:
• tRNAs bind to a particular amino acid, and
become “charged.”
• tRNAs bind at their midpoint—anticodon-to
mRNA molecules.
• tRNAs interacts with ribosomes.
Figure 10.13 Transfer RNA
Concept 10.4 Translation of the Genetic Code Is Mediated by
tRNA and Ribosomes
Wobble—specificity for the base at the 3′
end of the codon is not always observed.
Example: Codons for alanine—GCA, GCC,
and GCU—are recognized by the same
tRNA.
Wobble allows cells to produce fewer tRNA
species, but does not allow the genetic
code to be ambiguous.
In-Text Art, Ch. 10, p. 200
Concept 10.4 Translation of the Genetic Code Is Mediated by
tRNA and Ribosomes
Activating enzymes—aminoacyl-tRNA
synthetases—charge tRNA with the
correct amino acids.
Each enzyme is highly specific for one
amino acid and its corresponding tRNA.
The enzymes have three-part active sites—
they bind a specific amino acid, a specific
tRNA, and ATP.
Concept 10.4 Translation of the Genetic Code Is Mediated by
tRNA and Ribosomes
Experiment by Benzer and others:
Cysteine already bound to tRNA was
chemically changed to alanine.
Which would be recognized—the amino
acid or the tRNA in protein synthesis?
Answer: Protein synthesis machinery
recognizes the anticodon, not the amino
acid
Concept 10.4 Translation of the Genetic Code Is Mediated by
tRNA and Ribosomes
The translation of mRNA by tRNA is
accomplished at the ribosome—the
workbench—and holds mRNA and
charged tRNAs in the correct positions to
allow assembly of polypeptide chain.
Ribosomes are not specific; they can make
any type of protein.
Concept 10.4 Translation of the Genetic Code Is Mediated by
tRNA and Ribosomes
Ribosomes have two subunits, large and
small.
In eukaryotes, the large subunit has three
molecules of ribosomal RNA (rRNA) and
49 different proteins in a precise pattern.
The small subunit has one rRNA and 33
proteins.
Figure 10.14 Ribosome Structure
Concept 10.4 Translation of the Genetic Code Is Mediated by
tRNA and Ribosomes
Large subunit has three tRNA binding sites:
A (amino acid) site binds with anticodon of
charged tRNA.
P (polypeptide) site is where tRNA adds its
amino acid to the growing chain.
E (exit) site is where tRNA sits before being
released from the ribosome.
Concept 10.4 Translation of the Genetic Code Is Mediated by
tRNA and Ribosomes
Ribosome has a fidelity function: when
proper binding occurs, hydrogen bonds
form between the base pairs.
Small subunit rRNA validates the match—if
hydrogen bonds have not formed between
all three base pairs, the tRNA must be an
incorrect match for that codon and the
tRNA is rejected.
Concept 10.4 Translation of the Genetic Code Is Mediated by
tRNA and Ribosomes
Like transcription, translation also occurs in
three steps:
• Initiation
• Elongation
• Termination
Concept 10.4 Translation of the Genetic Code Is Mediated by
tRNA and Ribosomes
Initiation:
An initiation complex consists of a
charged tRNA and small ribosomal
subunit, both bound to mRNA.
After binding, the small subunit moves along
the mRNA until it reaches the start codon,
AUG.
The first amino acid is always methionine,
which may be removed after translation.
Concept 10.4 Translation of the Genetic Code Is Mediated by
tRNA and Ribosomes
The large subunit joins the complex; the
charged tRNA is now in the P site of the
large subunit.
Initiation factors are responsible for
assembly of the initiation complex from
mRNA, two ribosomal subunits and
charged tRNA.
Figure 10.15 The Initiation of Translation (Part 1)
Figure 10.15 The Initiation of Translation (Part 2)
Concept 10.4 Translation of the Genetic Code Is Mediated by
tRNA and Ribosomes
Elongation: The second charged tRNA
enters the A site
Large subunit catalyzes two reactions:
It breaks bond between tRNA in P site and
its amino acid.
A peptide bond forms between that amino
acid and the amino acid on tRNA in the A
site.
Concept 10.4 Translation of the Genetic Code Is Mediated by
tRNA and Ribosomes
When the first tRNA has released its
methionine, it moves to the E site and
dissociates from the ribosome—it can then
become charged again.
Elongation occurs as the steps are
repeated, assisted by proteins called
elongation factors.
Concept 10.4 Translation of the Genetic Code Is Mediated by
tRNA and Ribosomes
The large subunit has peptidyl transferase
activity—if rRNA is destroyed, the activity
stops.
The component with this activity is an rRNA
in the ribosome.
The catalyst is an example of a ribozyme
(from ribonucleic acid and enzyme).
Figure 10.16 The Elongation of Translation (Part 1)
Figure 10.16 The Elongation of Translation (Part 2)
Concept 10.4 Translation of the Genetic Code Is Mediated by
tRNA and Ribosomes
Termination—translation ends when a stop
codon enters the A site.
Stop codon binds a protein release factor—
allows hydrolysis of bond between
polypeptide chain and tRNA on the P site.
Polypeptide chain separates from the
ribosome—C terminus is the last amino
acid added.
Figure 10.17 The Termination of Translation (Part 1)
Figure 10.17 The Termination of Translation (Part 2)
Table 10.2 Signals that Start and Stop Transcription and Translation
Concept 10.4 Translation of the Genetic Code Is Mediated by
tRNA and Ribosomes
Several ribosomes can work together to
translate the same mRNA, producing
multiple copies of the polypeptide.
A strand of mRNA with associated
ribosomes is called a polyribosome, or
polysome.
Concept 10.5 Proteins Are Modified after Translation
Posttranslational aspects of protein
synthesis:
Polypeptide emerges from the ribosome and
folds into its 3-D shape.
Its conformation allows it to interact with
other molecules—it may contain a signal
sequence (or signal peptide) indicating
where in the cell it belongs.
Concept 10.5 Proteins Are Modified after Translation
In the absence of a signal sequence, the
protein will remain where it was made.
Some proteins contain signal sequences
that “target” them to the nucleus,
mitochondria, or other places.
Signal sequence binds to a receptor protein
on the organelle surface—a channel forms
and the protein moves into the organelle.
Figure 10.19 Destinations for Newly Translated Polypeptides in a Eukaryotic Cell (Part 1)
Figure 10.19 Destinations for Newly Translated Polypeptides in a Eukaryotic Cell (Part 2)
Figure 10.20 Testing the Signal
Concept 10.5 Proteins Are Modified after Translation
Protein modifications:
Proteolysis—cutting of a long polypeptide
chain, or polyprotein, into final products, by
proteases
Glycosylation—addition of carbohydrates
to form glycoproteins
Phosphorylation—addition of phosphate
groups catalyzed by protein kinases—
charged phosphate groups change the
conformation of the protein
Figure 10.21 Posttranslational Modifications of Proteins
Answer to Opening Question
Tetracyclines kill bacteria by interrupting
translation.
They bind to the small subunit of the
ribosome, which changes the ribosome
structure.
Charged tRNAs can no longer bind to the A
site on the ribosome.
Figure 10.22 An Antibiotic at the Ribosome